EP0468042B1 - HOMOLOGATED VITAMIN D2 COMPOUNDS AND THE CORRESPONDING 1alpha-HYDROXYLATED DERIVATIVES - Google Patents

HOMOLOGATED VITAMIN D2 COMPOUNDS AND THE CORRESPONDING 1alpha-HYDROXYLATED DERIVATIVES Download PDF

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EP0468042B1
EP0468042B1 EP91905084A EP91905084A EP0468042B1 EP 0468042 B1 EP0468042 B1 EP 0468042B1 EP 91905084 A EP91905084 A EP 91905084A EP 91905084 A EP91905084 A EP 91905084A EP 0468042 B1 EP0468042 B1 EP 0468042B1
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hydroxy
vitamin
solution
compound
mixture
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EP0468042A1 (en
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Hector Floyd Deluca
Heinrich Konstantine Schnoes
Kato Leonard Perlman
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Wisconsin Alumni Research Foundation
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07J—STEROIDS
    • C07J71/00—Steroids in which the cyclopenta(a)hydrophenanthrene skeleton is condensed with a heterocyclic ring
    • C07J71/0036—Nitrogen-containing hetero ring
    • C07J71/0042—Nitrogen only
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P5/00—Drugs for disorders of the endocrine system
    • A61P5/10—Drugs for disorders of the endocrine system of the posterior pituitary hormones, e.g. oxytocin, ADH
    • A61P5/12—Drugs for disorders of the endocrine system of the posterior pituitary hormones, e.g. oxytocin, ADH for decreasing, blocking or antagonising the activity of the posterior pituitary hormones
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C401/00—Irradiation products of cholesterol or its derivatives; Vitamin D derivatives, 9,10-seco cyclopenta[a]phenanthrene or analogues obtained by chemical preparation without irradiation

Definitions

  • This invention relates to biologically active vitamin D compounds.
  • this invention relates to novel 24, 26 and/or 27 homalogated derivatives of vitamin D2 and the corresponding hydroxylated forms thereof.
  • the D vitamins are very important agents for the control of calcium and phosphate metabolism in animals and humans, and have long been used as dietary supplements and in clinical practice to assure proper bone growth and development. It is now known that the in vivo activity of these vitamins, specifically of vitamin D2 and D3, is dependent on metabolism to hydroxylated forms. Thus, vitamin D3 undergoes two successive hydroxylation reactions in vivo, leading first to 25-hydroxyvitamin D3 and then to 1,25-dihydroxyvitamin D3 and the latter is thought to be the compound responsible for the well-known beneficial effects of vitamin D3.
  • vitamin D2 which is commonly used as a dietary supplement, undergoes an analogous hydroxylation sequence to its active forms, being first converted to 25-hydroxyvitamin D2 (25-OH-D2) and then to 1,25-dihydroxyvitamin D2 (1,25-(OH)2D2).
  • vitamin D2 Like the metabolites of the vitamin D3 series, the hydroxylated forms of vitamin D2 named above are, because of their potency and other beneficial properties, highly desirable dietary supplements, or pharmaceutical agents, for the cure or prevention of bone or related diseases, and their value and possible use is recognized in patents relating to these compounds [U. S. Letters Pat. Nos. 3,585,221 and 3,880,894].
  • These compounds likewise elicit the full spectrum of vitamin D-type responses such as intestinal calcium transport, bone mineral mobilization and bone calcification response in the animal or human.
  • 1 ⁇ ,25-dihydroxyvitamin D2 and 1 ⁇ -hydroxyvitamin D2 are characterized by having a C-24 stereochemistry as it occurs in the side chain of ergosterol, i.e. these compounds are defined by the structures shown below, where R represents side chains (a) and (b), respectively: More recently the C-24-epimer of 1 ⁇ ,25-dihydroxyvitamin D2 and 1 ⁇ -hydroxyvitamin D2 has been prepared and tested. These compounds are characterized by the structures shown above, where R represents side chains (c) and (d) respectively. Remarkably these C-24-epimeric vitamin D derivatives exhibit a distinctly different biological activity in that they promote intestinal calcium absorption and the calcification of bone, but elicit little or no bone calcium mobilization response.
  • This invention provides new homologated vitamin D2 analogs which may be represented by the structure below, as well as the 1 ⁇ -hydroxy-analogs and the protected hydroxy derivatives of the compounds: where the configuration about carbon 24 may be R or S and wherein n is an integer having a value of from 1 to 5, X1 is selected from hydrogen or a hydroxy protecting group, X2 is selected from hydrogen, hydroxy, and protected hydroxy, X3 is selected from hydrogen, hydroxy, and protected hydroxy, R3 is selected from alkyl, hydroxy, protected hydroxy, hydrogen or fluorine, and wherein R1 and R2, which may be the same or different, are each selected from an alkyl or aryl group, with the proviso that when X1 is hydrogen, X2 is hydrogen or hydroxy, X3 is hydrogen or hydroxy and n is 1, R1 and R2 cannot both be methyl and R3 cannot be hydrogen or methyl.
  • These compounds are distinguished from the previously known 1 ⁇ -hydroxyvitamin D2 and 1 ⁇ -hydroxy-24-epi-vitamin D2 compounds by homologation at the 24, 26 and/or 27 positions.
  • Specific compounds disclosed include 26,27-dihomo-1 ⁇ -hydroxyvitamin D2, 26,27-dihomo-1 ⁇ ,25-dihydroxyvitamin D2, 26,27-dihomo-1 ⁇ -hydroxy-24-epi-vitamin D2, 26,27-dihomo-1 ⁇ , 25-dihydroxy-24-epi-vitamin D2, and the corresponding 26,27-tetrahomo compounds, as well as 24-dihomo-1 ⁇ -hydroxy-24-epi-vitamin D2, 24-dihomo-1 ⁇ ,25-dihydroxy-24-epi-vitamin D2, and the corresponding 24-trihomo compounds.
  • the term “24-dihomo” refers to the addition of two methylene groups substituted with the group R3 at the carbon 24 position in the side chain (so that n is 3).
  • the term “24-trihomo” refers to the addition of three such substituted methylene groups (so that n is 4).
  • the term “26,27-dihomo” refers to the addition of a methyl group at the carbon 26 and 27 positions so that R1 and R2 are ethyl groups.
  • the term “26,27-tetrahomo” refers to the addition of an ethyl group at the 26 and 27 positions so that R1 and R2 are propyl groups.
  • vitamin D2 A novel and convenient synthesis of vitamin D2 compounds has now been developed and is also described herein.
  • This synthesis provides homologated vitamin D2 compounds characterized by the structures previously shown as well as non-homologated vitamin D2 compounds shown below where X2 is hydrogen, such as vitamin D2 itself and the 24-epimer of vitamin D2, namely 24-epi-vitamin D2 (24-epi D2), characterized by the structures shown below where X1 and X2 are both hydrogen and R1, R2 and R3 are methyl.
  • This synthesis also provides 25 hydroxylated vitamin D2 compounds, such as 25-hydroxyvitamin D2 (25-OH-D2) and the 24-epimer of 25-hydroxyvitamin D2, namely 25-hydroxy-24-epi-vitamin D2 (25-OH-24-epi D2), characterized by the structures shown below where X1 is hydrogen, X2 is hydroxy and R1, R2 and R3 are methyl.
  • X2 may be either hydrogen or hydroxy, as well as the corresponding alkyl or aryl analogs thereof, characterized by the structure above where R1 and R2 are alkyl or aryl, and the corresponding side chain substituted derivatives there R3 is alkyl, hydroxy, protected hydroxy (as defined below for X1 and X2), hydrogen or fluorine, and the hydroxy-protected derivatives of these compounds characterized by the structures above, where X1 is selected from the group consisting of acyl, alkylsilyl, or alkoxyalkyl and X2 is selected from the group consisting of O-acyl, O-alkylsilyl, or O-alkoxyalkyl.
  • the present process provides the 5,6-trans-isomers of the above compounds, as well as the 24 and/or 26 and/or 27 homologated compounds previously shown herein.
  • the above compounds can be 1 ⁇ -hydroxylated by known methods, so as to produce the corresponding 1 ⁇ -hydroxyvitamin D derivatives.
  • Especially preferred examples of the latter are 1 ⁇ -hydroxyvitamin D2, 1 ⁇ ,25-dihydroxyvitamin D2, 1 ⁇ -24-epi-vitamin D2 and 1 ⁇ ,25-dihydroxy-24-epi-vitamin D2.
  • acyl signifies an aliphatic acyl group of from 1 to about 6 carbons, in all possible isomeric forms (e.g. formyl, acetyl, propionyl, butyryl, isobutyryl, valeryl, etc.), or an aromatic acyl group (aroyl group) such as benzoyl, the isomeric methtlbenzoyls, the isomeric nitro- or halo-benzoyls, etc., or a dicarboxylic acyl group of from 2 to 6 atoms chain length, i.e.
  • alkyl refers to a lower alkyl group of 1 to 6 carbons in all possible isomeric forms, e.g.
  • alkylsilyl refers to trialkyl silicone groupings where the alkyl groups may be the same or different as exemplified by trimethylsilyl, triethylsilyl, dimethyl-tert.-butylsilyl and similar groupings.
  • alkoxyalkyl refers to protecting groups such as methoxymethyl, ethoxymethyl, methoxyethoxymethyl, and similar alkoxymethyl groups, as well as the related cyclic structures, such as tetrahydropyranyl or tetrahydrofuranyl.
  • the overall process developed for the preparation of the above compounds may be divided into two general phases, namely (a) addition of a completed, preformed side chain fragment to a suitable steroidal precursor to produce a 5,7-diene steroid as the central intermediate, and (b) conversion of this 5,7-diene to the vitamin D structure to produce the desired vitamin D compound, and, if desired, (c) further conversion of the latter product to the corresponding 1 ⁇ -hydroxyvitamin D compound.
  • This process avoids the relatively difficult step of isomer separation which is required after conversion in the process of U. S. patent No. 4,448,721.
  • the process of the present invention also increases the yield of the end product since it utilizes a completed, preformed pure isomer side chain fragment to make the desired end product, rather than a mixture of side chain isomers as in U. S. Patent No. 4,448,721.
  • the process of this invention comprises the reaction of a steroidal 22-aldehyde of the structure: where X1 is hydrogen or a hydroxy-protecting group, with a sulfone derivative of the general formula, ArSO2CH2R where Ar represents a phenyl or tolyl group, and R is selected from the group consisting of straight or branched, substituted or unsubstituted, hydrocarbon radicals of from 1 to 25 carbon atoms, where the substituents are selected from the group consisting of hydroxy, protected hydroxy, and fluorine.
  • the coupling reaction conducted in a basic medium, between the above aldehyde and sulfone derivatives, yields a condensation product of the formula: which is then subjected to reduction (either as the 22-hydroxy, or as the corresponding 22-O-acylated derivative) using metal amalgams (Na, Al, Zn amalgams) or related dissolving metal reduction systems, so as to provide the 22, 23-unsaturated steroid of the formula: where R and X1 represent the groupings defined above.
  • This steroid intermediate can then be further converted by known reactions to the desired vitamin D compounds.
  • Process Scheme I presents a specific embodiment of the overall process
  • Process Scheme II illustrates preparation of an appropriate side chain unit for addition to the steroid-22-aldehyde as shown in Scheme I.
  • steroidal 22-aldehydes such as, for example, the PTAD-diene-protected-22-aldehyde (4) shown in Scheme I (where PTAD refers to the phenyltriazoline-3,5-dione-protecting group shown), which in turn can be prepared from ergosterol by the known steps (Scheme I).
  • the first step of this process comprises the addition of a suitable side chain fragment.
  • a suitable side chain fragment For example, condensation of aldehyde (4) with a sulfonyl-side chain fragment as shown in Scheme I (sulfone (21), further described below) present in the form of its anion, in an ether or hydrocarbon solvent, provides the hydroxy-sulfone intermediate (5).
  • the anion of the sulfone (21) side chain fragment is generated by treatment of the sulfone with a strong base, such as lithium diethylamide, n-butyl lithium or methyl or ethyl magnesium bromide (or similar Grignard reagent) in an ether or hydrocarbon solvent, and to this solution of sulfone anion the steroid aldehyde (compound 4) as an ether or hydrocarbon solution is then added.
  • a strong base such as lithium diethylamide, n-butyl lithium or methyl or ethyl magnesium bromide (or similar Grignard reagent) in an ether or hydrocarbon solvent
  • the next step comprises the removal of the hydroxy-and phenylsulfonyl groups in the side chain with formation of the 22(23)-trans-double bond.
  • treatment of compound (5), in methanol solution saturated with NaHPO4, with sodium amalgam under an inert atmosphere gives compound (6) featuring the desired trans-22-double bond in the side chain.
  • the 22-hydroxy group in compound (5) can also be acylated or sulfonylated (e.g. mesylated) prior to the Na/Hg-reduction step, but this is not generally required.
  • the next operation of the process involves conversion of these ring B-protected steroids to the desired 5,7-diene intermediate (7).
  • this conversion is accomplished in a single step, namely treatment of (6) with a strong hydride reducing agent (e.g. LiAlH4) in an ether solvent at reflux temperature gives the diene (7).
  • a strong hydride reducing agent e.g. LiAlH4
  • the diene (7) is then converted to its 25-hydroxylated form (8) by known procedures in accordance with Scheme I.
  • Conversion of 5,7-diene (8) to the final vitamin D products (10) or (15) comprises a sequence of several steps.
  • the sequence shown in Process Scheme I involves first the irradiation of an ether or hydrocarbon solution of the 5,7-diene (8) with ultraviolet light to yield the previtamin analog (9) which by warming (50°-90°C) in a suitable solvent (e.g. ethanol, hexane) undergoes isomerization to the 25-hydroxyvitamin D2 compound (10).
  • a suitable solvent e.g. ethanol, hexane
  • compound (10) may be converted to the 1,25-dihydroxyvitamin D2 compound (15) by the known steps shown in Scheme I.
  • the side chain fragment, sulfone (21), as used in Scheme I is specifically the ( R ) enantiomer. Therefore, compounds (10) or (15) are obtained as the C-24-R-epimers, 25-hydroxy-24-epi-vitamin D2 (10) or 1,25-dihydroxy-24-epi-vitamin D2 (15), respectively.
  • compound (10) or (15) are prepared in epimerically pure form, and C-24-epimer separation as required in the process disclosed in U. S. Patent No. 4,448,721, is not necessary.
  • Use of the ( S )-epimer of sulfone (21) in the present process yields specifically 25-OH-D2, as well as, of course, the respective 1,25-dihydroxyvitamin D2 compound.
  • the 5,7-diene (7) may be used as the free hydroxy compound or as its hydroxy-protected form, where the hydroxy-protecting groups (at C-3 and/or C-25) may be acyl, alkylsilyl or alkoxyalkyl groups as previously defined.
  • the 25-OH-D2 product will be obtained as the free hydroxy compound or, if desired, as the C-3, or C-25-hydroxy-protected, or 3,25-dihydroxy-protected derivatives.
  • 25-OH-D2 epimers i.e. 25-OH-D2 or 25-OH-24-epi-D2 (10) when obtained in the free hydroxy forms, are also conveniently hydroxy-protected at the C-3 or C-25, or at both positions, by conventional reactions known in the art.
  • 25-OH-D2 may be acylated to yield, for example, the 25-OH-D2-3-acetate, or the corresponding 3,25-diacetate.
  • the 3-monoacetate in a like fashion, may be further acylated at C-25 by treatment with a different acylating reagent, or, alternatively, the 3,25-diacetate may be selectively hydrolyzed by a mild base (KOH/MeOH) to give the 25-monoacetate, which if desired can be reacylated with a different acyl group at C-3.
  • KOH/MeOH a mild base
  • Other hydroxy-protecting groups can be introduced by analogous known reactions.
  • the 5,6-trans-isomers of 25-OH-24-epi-D2 as well as the 1,25-dihydroxy compounds are compounds of potential utility in medical applications because of their considerable vitamin D-like activity.
  • These 5,6-trans-compounds are prepared from the 5,6-cis-isomers (i.e. 10 or 15) by iodine catalyzed isomerization according to the procedures of Verloop et al. Rec. Trav. Chim.
  • the required side chain fragment, sulfone (21), is itself prepared according to the process shown in Process Scheme II.
  • This synthesis is straight-forward and involves as a first step the reaction of ester (16) in anhydrous tetrahydrofuran (THF) with methyl magnesium bromide to give the diol (17).
  • Diol (17) is dissolved in anhydrous pyridine and reacted with p-toluenesulfonyl chloride to provide the tosylate (18).
  • Tosylate (18) is dissolved in a solution of anhydrous dimethyl formamide and reacted with thiophenol and t-BuOK to yield the sulfide (19).
  • the sulfide (19) in turn is dissolved in dichloromethane and reacted with 3-chloroperoxybenzoic acid to give the hydroxy sulfone compound (20).
  • Pyridinium p-toluenesulfonate is then added to a solution of compound (20) in anhydrous dichloromethane and reacted with dihydropyran to yield the hydroxy protected tetrahydropyranyl sulfone (21a).
  • the corresponding ( S )-epimer of sulfone (21) is prepared by the same process, using as starting material the ester corresponding to (16) but having the ( S ) configuration at carbon-2.
  • Process Scheme III presents another specific embodiment of the overall process, whereas Process Scheme IV illustrates preparation of an appropriate side chain unit for addition to the steroid-22-aldehyde as shown in Scheme III.
  • steroidal 22-aldehydes such as, for example, the PTAD-diene-protected-22-aldehyde (4) shown in Scheme I (where PTAD refers to the phenyltriazoline-3,5-dione-protecting group shown), which in turn can be prepared from ergosterol by the known steps (Scheme I).
  • the first step of this process comprises the addition of a suitable side chain fragment.
  • a suitable side chain fragment For example, condensation of aldehyde (4) with a sulfonyl-side chain fragment as shown in the Scheme III (sulfone (35), further described below) present in the form of its anion, in an ether or hydrocarbon solvent, provides the hydroxy-sulfone intermediate (22).
  • the anion of the sulfone (35) side chain fragment is generated by treatment of the sulfone with a strong base, such as lithium diethylamide, n-butyl lithium or methyl or ethyl magnesium bromide (or similar Grignard reagent) in an ether or hydrocarbon solvent, and to this solution of sulfone anion the steroid aldehyde (compound 4) as an ether or hydrocarbon solution is then added.
  • a strong base such as lithium diethylamide, n-butyl lithium or methyl or ethyl magnesium bromide (or similar Grignard reagent) in an ether or hydrocarbon solvent
  • the next step comprises the removal of the hydroxy-and phenylsulfonyl groups in the side chain with formation of the 22(23)-trans-double bond.
  • treatment of compound (22), in methanol solution saturated with NaHPO4, with sodium amalgam under an inert atmosphere gives compound (23) featuring the desired trans-22-double bond in the side chain.
  • the 22-hydroxy group in compound (22) can also be acylated or sulfonylated (e.g. mesylated) prior to the Na/Hg-reduction step, but this is not generally required.
  • the next operation of the process involves conversion of these ring B-protected steroids to the desired 5,7-diene intermediate (24).
  • this conversion is accomplished in a single step, namely treatment of (23) with a strong hydride reducing agent (e.g. LiAlH4) in an ether solvent at reflux temperature gives the diene (24).
  • a strong hydride reducing agent e.g. LiAlH4
  • Conversion of 5,7-diene (24) to the final vitamin D products (26) or (31) comprises a sequence of several steps.
  • the sequence shown in Process Scheme III involves first the irradiation of an ether or hydrocarbon solution of the 5,7-diene (24) with ultraviolet light to yield the previtamin analog (25) which by warming (50°-90°C) in a suitable solvent (e.g. ethanol, hexane) undergoes isomerization to the vitamin D2 compound (26).
  • a suitable solvent e.g. ethanol, hexane
  • compound (26) may be converted to the 1 ⁇ -hydroxyvitamin D2 compound (31) by the known steps shown in Scheme III.
  • the side chain fragment, sulfone (35), as used in Scheme III is specifically the ( R ) enantiomer. Therefore, compounds (26) or (31) are obtained as the C-24-R-epimers, 24-epi-vitamin D2 (26) or 1 ⁇ -hydroxy-24-epi-vitamin D2 (31), respectively. Thus, compounds (26) or (31) are prepared in epimerically pure form, and C-24-epimer separation as required in the process disclosed in U. S. patent No. 4,448,721, is not necessary. Use of the ( S )-epimer of sulfone (35) in the present process yields specifically vitamin D2, as well as, of course, the respective 1 ⁇ -hydroxyvitamin D2 compound.
  • the 5,7-diene (24) may be used as the free hydroxy compound or as its hydroxy-protected form, where the hydroxy-protecting groups (at C-3) may be acyl, alkylsilyl or alkoxyalkyl groups as previously defined.
  • the vitamin D2 product will be obtained as the free hydroxy compound or, if desired, as the C-3-hydroxy-protected derivatives.
  • Synthesis according to Scheme III would provide the vitamin D2 products as the free hydroxy compounds but analogous conversion of 5,7-diene intermediate (24) as the 3-protected, derivative will yield the corresponding hydroxy-protected derivatives of the vitamin D2 products.
  • vitamin D2 epimers i.e. vitamin D2 or 24-epi-D2 (26) when obtained in the free hydroxy forms, are also conveniently hydroxy-protected at the C-3 position, by conventional reactions known in the art.
  • vitamin D2 may be acylated to yield, for example, the vitamin D2-3-acetate.
  • Other hydroxy-protecting groups can be introduced by analogous known reactions.
  • the 5,6-trans-isomers of 24-epi-D2 as well as the 1 ⁇ -hydroxy compounds are compounds of potential utility in medical applications because of their considerable vitamin D-like activity.
  • These 5,6-trans-compounds are prepared from the 5,6-cis-isomers (i.e. 26 or 31) by iodine catalyzed isomerization according to the procedures of Verloop et al. Rec. Trav. Chim. Pays Bas 78, 1004 (1969), and the corresponding 3-hydroxy-protected derivatives are likewise obtained by analogous isomerization of the corresponding 5,6-cis-acylates, or by hydroxy-protection of the 5,6-trans-D2 compounds.
  • the required side chain fragment, sulfone (35), can be prepared according to Perlman et al, supra , or according to the process shown in Process Scheme IV.
  • This synthesis is straightforward and involves as a first step dissolving alcohol (32) in anhydrous pyridine and reacting it with p-toluenesulfonyl chloride to provide the tosylate (33).
  • Tosylate (33) is dissolved in a solution of anhydrous dimethyl formamide and reacted with thiophenol and t-BuOK to yield the sulfide (34).
  • the sulfide (34) in turn is dissolved in dichloromethane and reacted with 3-chloroperoxybenzoic acid to give the sulfone compound (35).
  • the corresponding ( S )-epimer of sulfone (35) can also be prepared according to Perlman et al supra , or according to Process Scheme IV, using as starting material the alcohol corresponding to (32) but having the ( S ) configuration at carbon-2.
  • the present process also serves as a convenient method for the synthesis of side chain homologated vitamin D2 analogs where the carbon at any one of the side chain positions designated as 24, 26 and 27 may be homologated, of the formula (40) shown below, or of the corresponding 25-hydroxy-analogs and/or 1 ⁇ -hydroxy-analogs, where n is an integer having a value of from 1 to 5, X1 is selected from hydrogen and a hydroxy-protecting group, X2 is selected from hydrogen, hydroxy and protected hydroxy, X3 is selected from hydrogen, hydroxy, and protected hydroxy, R3 is alkyl, hydroxy, protected hydroxy, hydrogen or fluorine, and where R1 and R2, which may be the same or different, is an alkyl group or an aryl group and where the configuration about carbon 24 has either the ( R )- or the ( S )-stereochemical orientation.
  • These compounds are prepared by condensing compound (4) with the appropriate alkyl or aryl side chain fragment as shown by the following formula
  • the solution was cooled at -78°C and treated with an ozone-oxygen mixture for 30 min (TLC control) and thoroughly purged with nitrogen. 50ml of dimethyl sulfide was added and the mixture was washed with 300ml of water, 200ml of 2N HCl (twice) and 300ml of water. The organic layer was separated and each washing was extracted with 400ml and 200ml of chloroform. The combined extract was dried over Na2SO4 and concentrated in vacuo. The residue was purfied on a silica gel column (5.06X63cm, 550g of 150 ⁇ 425»m silica gel) using a mixture of ethyl acetate and hexane as eluant.
  • the mixture was stirred at -78°C for 1hr, decomposed by the addition of 100ml of saturated NH4Cl solution, warmed to 0°C and extracted three times with 100ml of ethyl acetate. Each extract was washed with 100ml of saturated NaCl solution, dried over Na2SO4 and concentrated in vacuo. The residue was purified on a silica gel column (3.2X60cm, 150g of 75 ⁇ 150»m silica gel). The unreacted sulfone 21 was eluted with benzene and 14.7g (92%) of 5 was eluted with ethyl acetate.
  • the residue was purified on a silica gel column (2.3X8.0cm, 10g of 45 ⁇ 75 »m silica gel) using a mixture of ethyl acetate in hexane as eluant. 310 mg of 15 was eluted with 30% ethyl acetate in hexane, combined with another 337mg of 15 and recrystallized from methyl formate.
  • a mixture of hydroxysulfones 22 , 5% sodium amalgum and methanol saturated with Na2HPO4 is stirred under nitrogen atmosphere.
  • the reaction solution is decanted and concentrated in vacuo.
  • the residue is dissolved in ethyl acetate and washed with water.
  • the ethyl acetate extract is separated and each washing is extracted twice with ethyl acetate.
  • the combined extract is dried over Na2SO4 and concentrated in vacuo to obtain 23 .
  • Compound 24 is dispersed in a mixture of ether and benzene (4:1) and irradiated with stirring under nitrogen in a water-cooled quartz immersion well equipped with an ozone free filter using a high-pressure UV lamp. The reaction may be monitored by HPLC.
  • the solution is concentrated in vacuo, redissolved in ethanol and heated under reflux and nitrogen. Then, the solution is concentrated in vacuo and the residue is purified on a silica gel column to obtain compound 26 .
  • Tert-butyl hydroperoxide (3.0M in 2,2-4-trimethylpentane) is added to a suspension of selenium dioxide in anhydrous dichloromethane. The mixture is stirred at room temperature under nitrogen. Anhydrous pyridine is added followed by a solution of compound 28 in anhydrous dichloromethane. The mixture is stirred under nitrogen at room temperature and heated under reflux. Then, a 10% NaOH solution is added and the mixture is extracted with ether. Each extract is washed with a 10% NaOH solution and a saturated NaCl solution. The combined extract is dried over Na2SO4 and concentrated in vacuo. The residue is purified on a silica gel column to obtain compound 29 .
  • Compound 29 is dissolved in acetic acid and heated under nitrogen. Then, the solution is poured over ice and neutralized with saturated NaHCO3 solution. The mixture is extracted three times with a mixture of ether and dichloromethane (4:1). Each extract is washed with a saturated NaHCO3 solution and a saturated NaCl solution. The combined extract is dried over Na2SO4 and concentrated in vacuo. Then, to a solution of the residue in ethyl acetate is added maleic anhydride and the mixture is allowed to stand under nitrogen at room temperature. Then, the solution is concentrated in vacuo, the residue is redissolved in ether. KOH in methanol is added, the solution is stirred at room temperature and concentrated in vacuo.
  • the residue is dissolved in a mixture of ether and dichloromethane (4:1) and washed with 10% NaOH solution twice and a saturated NaCl solution.
  • the organic extract is separated and each washing is extracted twice with the same mixture of ether and dichloromethane.
  • the combined extract is dried over Na2SO4 and concentrated in vacuo.
  • the residue is purified on a silica gel column using a mixture of ethyl acetate in hexane as eluant to obtain compound 31 .
  • the compounds of the present invention may be prepared in accordance with the procedures set forth in U. S. Patent No. 4,847,012. The following illustrates the preparation of 24-epi-26,27-dihomo 1 ⁇ ,25-dihydroxyvitamin D2. This compound is distinguished from the previously known 24-epi-1 ⁇ ,25-dihydroxyvitamin D2 by homologation at the 26,27 positions. As is readily apparent to those skilled in this art, the following described synthesis may be readily adapted to making the other homologated compounds disclosed herein by selecting the appropriate side chain unit and/or vitamin D nucleus for the condensation step.
  • the synthesis of 24-epi-26,27-dihomo-1 ⁇ ,25-dihydroxyvitamin D2 requires the construction of an appropriate side chain unit having the desired (S) stereochemistry at the carbon center that is to become carbon-(24 R) in the target compound, and the condensation of the side chain unit with a suitable 1 ⁇ -hydroxylated vitamin D nucleus so as to generate the desired final product.
  • the synthesis of the optically active side chain unit comprised the conversion of the commercially available (R)-(-)-3-hydroxy-2-methylpropionate 41 with ethylmagnesium bromide to the diol 42 .
  • This was converted to tosylate 43 , then treated with potassium thiophenoxide in alkaline DMF to give sulfide 44 .
  • This was oxidized with 3-chloroperbenzoic acid to afford sulfone 45 .
  • the very hindered hydroxy of sulfone 45 could not be protected by standard methodology. However, it was successfully protected by the use of triethylsilyltriflate in triethylamine to give the protected sulfone 46 .
  • the crude diol 42 (3 g)(20.5 mmol) was dissolved in 8 mL of anhydrous pyridine and 4.7 g (24.7 mmol) of p-toluenesulfonyl chloride was added with stirring at 0°C. The mixture was kept for 16 h at 4°C and quenched with ice water. The water phase was extracted with ether and the ether phase was washed with ice cold 1 N. HC1, saturated CuSO4 solution, water, NaHCO3 solution, brine and dried over anh. MgSO4, filtered and evaporated. The crude oil was purified by silica gel column chromatography with ethyl acetate - hexane mixtures to give 4.4 g (73%) tosylate 43 as a colorless oil.
  • Table 5 illustrates the cell differentiation activity of 26,27-dihomo-24-epi-1 ⁇ -hydroxyvitamin D2 and 26,27-dihomo-1 ⁇ -hydroxyvitamin D2 as compared with 26,27-dihomo-24-epi-1,25-dihydroxyvitamin D3.
  • the NBT (nitroblue tetrazolium reduction) and phago (phagocytosis assay) differentiation data of HL-60 cells were obtained from tessts conducted according to standard procedures (see e.g. U. S. Patent No. 4,973,584).
  • Rats were fed a 0.02% Ca, 0.3% P diet for 3 weeks and then given the indicated dose (100 pmol) intravenously or (325 pmol) intraperitoneally. The measurements were made 12.5 hours after the dose. There were 6 rats per group. TABLE 3 Calcium Transport and Bone Calcium Mobilization of Rats Chronically Dosed With 26,27-Dihomo-24-Epi-1 ⁇ -OH-D2 or 26,27-Dihomo-1 ⁇ -OH-D2 Compound Dose (pmoles/day) Ca Transport (S/M) Serum Calcium (Bone Ca Mobil.) (mg%) -D (Control 0 2.7 ⁇ 0.25 4.5 ⁇ 0.04 1 ⁇ -OH-D2 50 4.7 ⁇ 0.15 5.1 ⁇ 0.22 125 5.3 ⁇ 0.44 5.7 ⁇ 0 07 26,27-Dihomo-24-epi-1 ⁇ -OH-D2 50 -- 3.7 ⁇ 0.13 125 3.2 ⁇ 0.05 ⁇ 0.30 26,27-Dihomo-1 ⁇
  • Rats were fed a 0.02% calcium, 0.3% phosphorus vitamin D-deficient diet for 3 weeks and then given the indicated dose intraperitoneally in 0.05 ml of 95% ethanol each day for 7 days. The measurements were made 24 hours after the last dose. There were 6 rats in each group and the data are presented as the mean ⁇ SEM.
  • the rats were fed for 3 weeks on a high calcium (1.2%), low phosphorus (0.1%) diet, and then given the indicated dose in 0.05 ml 95% ethanol each day for 7 days. The measurements were made 24 hours after the last dose.
  • the preceding assays demonstrate that the new compounds, 26,27-dihomo-1 ⁇ -hydroxyvitamin D2 and 26,27-dihomo-1 ⁇ -hydroxy-24-epi-vitamin D2, exhibit a very distinct and unique spectrum of activities.
  • the 26,27-homologated epi-analog actually diminishes percent bone ash and total bone ash, indicating that it is antagonizing any remaining endogenous vitamin D found in the animals.
  • the epi-compound diminishes serum calcium below control values in the case of the animals maintained on a low calcium diet.
  • the novel compound of this invention may be formulated as a solution in innocuous solvents, or as an emulsion, suspension or dispersion in suitable solvents or carriers, or as pills, tablets or capsules, together with solid carriers, according to conventional methods known in the art.
  • the compounds are advantageously administered by injection, or by intravenous infusion of suitable sterile solutions, or in the form of liquid or solid doses via the alimentary canal.
  • Doses of from 1 »g to 50 »g per day, particularly of 1 ⁇ -hydroxy-24-epi-vitamin D2 are appropriate for treatment purposes, such doses being adjusted according to the disease to be treated and the response of the subject as is well understood in the art.
  • the new epi compound exhibits specificity of action, it is suitably administered alone, in situations where only calcium transport stimulation is desired, or together with graded doses of another active vitamin D compound -- e.g. 1 ⁇ -hydroxyvitamin D2 or D3, or 1 ⁇ ,25-dihydroxyvitamin D3 -- in situations where some degree of bone mineral mobilization (together with calcium transport stimulation) is found to be advantageous.
  • another active vitamin D compound e.g. 1 ⁇ -hydroxyvitamin D2 or D3, or 1 ⁇ ,25-dihydroxyvitamin D3 -- in situations where some degree of bone mineral mobilization (together with calcium transport stimulation) is found to be advantageous.

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EP91905084A 1990-02-14 1991-02-13 HOMOLOGATED VITAMIN D2 COMPOUNDS AND THE CORRESPONDING 1alpha-HYDROXYLATED DERIVATIVES Expired - Lifetime EP0468042B1 (en)

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AU656829B2 (en) * 1991-12-26 1995-02-16 Wisconsin Alumni Research Foundation 26, 27-dimethylene-1 alpha, 25-dihydroxyvitamin D2 and 26,27-dihydroxyvitamin D2 and methods for preparing same
US5716946A (en) * 1996-02-13 1998-02-10 Wisconsin Alumni Research Foundation Multiple sclerosis treatment
ATE246172T1 (de) * 1998-05-29 2003-08-15 Bone Care Int Inc Verfahren zur herstellung von hydroxy-25-ene vitamin d verbindungen
US6479474B2 (en) * 1999-07-08 2002-11-12 Wisconsin Alumni Research Foundation Dietary calcium as a supplement to vitamin D compound treatment of multiple sclerosis
US7094775B2 (en) 2004-06-30 2006-08-22 Bone Care International, Llc Method of treating breast cancer using a combination of vitamin D analogues and other agents
US20060003950A1 (en) * 2004-06-30 2006-01-05 Bone Care International, Inc. Method of treating prostatic diseases using a combination of vitamin D analogues and other agents
EP1812011A1 (en) 2004-11-12 2007-08-01 Bioxell S.p.a. Combined use of vitamin d derivatives and anti-proliferative agents for treating bladder cancer
US7745226B2 (en) 2005-04-06 2010-06-29 Quest Diagnostics Investments Incorporated Methods for detecting vitamin D metabolites
US7972868B2 (en) 2007-11-28 2011-07-05 Quest Diagnostics Investments Incorporated Methods for detecting dihydroxyvitamin D metabolites by mass spectrometry
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EP3603646B1 (en) 2009-08-14 2024-04-03 BPGbio, Inc. Vitamin d3 and analogs thereof for treating alopecia
US7977117B2 (en) 2009-12-03 2011-07-12 Quest Diagnostics Investments Incorprated Vitamin D metabolite determination utilizing mass spectrometry following derivatization
CN107607662B (zh) 2009-12-11 2020-05-08 奎斯特诊断投资公司 多重样品中的甾族化合物的质谱法
CN102753964A (zh) 2009-12-11 2012-10-24 奎斯特诊断投资公司 非衍生的、非代谢的维生素d的质谱测定
CN112156097A (zh) 2013-05-29 2021-01-01 博格有限责任公司 使用维生素d预防或减轻化疗诱发的脱发

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JP2731839B2 (ja) 1998-03-25
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DE69109006D1 (de) 1995-05-24
ATE121387T1 (de) 1995-05-15

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